PCRを用いた遺伝子変異の迅速検出法 : Sequential Multiplex Primers(SMP)法の基礎的検討

概要

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The authors have developed a simple and fast method called SMP-PCR to search for point mutations in a genome. The principle of this SMP-PCR depends on the forming DNA-ladder and their lacks by the multiplex primers in a tube of PCR. The primer containing mutation cannot be effective as a primer of PCR. So, PCR products have disappeared. For scanning point mutations in an enterotoxigenic Escherichia coil (ETEC) heat-labile toxin (LT) gene, eleven 13 mer-primers as forward-primers and a common reverseprimer were prepared. 3 typical strains, a wild and two mutant strain 35 and 43, were analyzed as models of this detection method. The target region (183 bp) of the LT-gene was amplified previously by first PCR, and then SMP-PCR was developed by using five different forward-primers, a common reverse-primer and 1st. PCR-product. On the optimum conditions of both concentration of primers and annealing-temperature, the difference in electrophoretic patterns of PCR products after SMP-PCR among a wild and two mutant strains have successed to detect. It is possible the sites of mutations are restricted within 10 bases by this method.
神戸大学の論文